Rneasy rna purification kit

Manufactured by Qiagen

Sourced in Germany, United States, United Kingdom

The RNeasy RNA purification kit is a laboratory equipment designed to isolate and purify RNA from a variety of sample types. It utilizes a silica-membrane-based technology to efficiently capture and elute RNA molecules.

Automatically generated - may contain errors

Lab products found in correlation

Close spelling variants of this product

RNeasy kit (11240 citations) RNeasy purification kit (154 citations) RNA purification kit (91 citations) RNeasy Mini Total RNA Purification kits (3 citations)

73 protocols using rneasy rna purification kit

Site-Directed Mutagenesis of rat nAChR α7

Cited 1 time

Check if the same lab product or an alternative is used in the 5 most similar protocols

The QuikChange protocol (Stratagene) was used for site-directed mutagenesis of the rat nAChR α7 subunit (pAMV vector). NotI was used to linearize the circular plasmid. The DNA was purified (Qiagen) and in vitro transcription of mRNA from the linearized DNA templates was performed using the T7 mMessage Machine kit (Ambion). The resulting mRNA was purified and isolated using Qiagen’s RNeasy RNA purification kit. The same linearization and mRNA synthesis protocols were used for the human Ric3 (pAMV) accessory protein.
For non-canonical amino acid incorporation, the amber (UAG) stop codon was used for all α7 subunit incorporation. The 74-nucleotide THG73 tRNA and 76-nucleotide THG73 tRNA were in vitro transcribed using the MEGAshortscript T7 (Ambion) kit and isolated using Chroma Spin DEPC-H₂O columns (Clontech). Synthesized non-canonical amino acids coupled to the dinucleotide dCA were enzymatically ligated to the 74-nucleotide tRNA as previously described (Nowak, et al., 1998 (link); Xiu, et al., 2009 (link)).
ND96 media was used for all experimental running/wash buffers (96 mM NaCl, 1.8mM CaCl₂, 2 mM KCl, 1 mM MgCl₂, 5 mM HEPES at pH 7.5). ND96+ media was used for oocyte storage media (2.5mM sodium pyruvate and 6.7mM theophylline added). No gentamicin was added to the ND96+ storage media in order to avoid distorting ACh-induced responses (Amici, et al., 2005 (link)).

Marotta C.B., Lester H.A, & Dougherty D.A. (2015). An unaltered orthosteric site and a network of long-range allosteric interactions for PNU-120596 in α7 nicotinic acetylcholine receptors. Chemistry & biology, 22(8), 1063-1073.

+ Open protocol

+ Expand

Transcriptional Response of MDA-MB-231 Cells to Cisplatin and TM

Check if the same lab product or an alternative is used in the 5 most similar protocols

1.0 × 10⁵ basal type MDA-MB-231 breast cancer cells (ATCC) were treated with 36 μM cisplatin (Sigma), 10 μM TM, both drugs, or a vehicle control in triplicate in 6-well plates and harvested at 12 and 24 hours. RNA was prepared from cell pellets using the Qiagen RNeasy RNA purification kit. RNA quality was determined by 2100 Bioanalyzer analysis (Agilent Technologies) and microarray was performed in triplicate for each drug treatment group using Affymetrix gene Chip Human Gene 1.0 ST Arrays (901086). ANOVA and pathway analysis was performed by the NIDDK Genomic Core Facility. The microarray data have been submitted to the GEO database under the accession number GSE77515.

Chisholm C.L., Wang H., Wong A.H., Vazquez-Ortiz G., Chen W., Xu X, & Deng C.X. (2016). Ammonium tetrathiomolybdate treatment targets the copper transporter ATP7A and enhances sensitivity of breast cancer to cisplatin. Oncotarget, 7(51), 84439-84452.

+ Open protocol

+ Expand

RNA Extraction and Quantification

Check if the same lab product or an alternative is used in the 5 most similar protocols

RNA was extracted using the RNeasy RNA Purification Kit (QIAGEN, cat #:
74106). Purified RNA was reverse transcribed using qScript cDNA synthesis kit
(Quanta Biosciences, cat #: 95047). Real-time PCR was performed in
triplicate using iQ SYBR Green-based detection on a ABI 7900HT fast qPCR
machine. The derived mRNA levels were normalized using RpLp0 mRNA levels. For
primer sequences, see Supplementary
Table 1.

Lawless S.J., Kedia-Mehta N., Walls J.F., McGarrigle R., Convery O., Sinclair L.V., Navarro M.N., Murray J, & Finlay D.K. (2017). Glucose represses dendritic cell-induced T cell responses. Nature Communications, 8, 15620.

+ Open protocol

+ Expand

Quantitative Analysis of Thyroid Genes

Check if the same lab product or an alternative is used in the 5 most similar protocols

Total RNA was extracted from slices using a Qiagen RNeasy RNA purification kit. cDNA was synthesized from 500 ng total RNA using High Capacity RNA‐to‐cDNA Master Mix (Applied Biosystems Ltd). Dio2 and Dio3 primers were obtained from Qiagen (QuantiTect Primer Assays Rn_Dio2_2_SG and Rn_Dio3_1_SG, respectively); other primers (Table 1) were designed using PrimerBLAST (Ye et al., 2012). qPCR reactions were set up using SensiMix SYBR master mix (Bioline) and were run on a Roche LightCycler 480 and analysed using LightCycler 480 1.5 software. Expression of genes of interest was normalized to Actb levels. Standard curves and blank controls were run for all sets of primers. Results shown are from a minimum of two independent experiments per condition. Gene expression in T3‐injected rats was compared to that of vehicle‐injected controls using unpaired Student's t‐tests. Expression in treated hypothalamic slices was compared to control slices from the same individuals by paired Student's t‐tests or ANOVA.

Stoney P.N., Helfer G., Rodrigues D., Morgan P.J, & McCaffery P. (2015). Thyroid hormone activation of retinoic acid synthesis in hypothalamic tanycytes. Glia, 64(3), 425-439.

+ Open protocol

+ Expand

Gene Expression Analysis of Cultured Cells

Check if the same lab product or an alternative is used in the 5 most similar protocols

RNA from cultured cells was isolated and purified using the RNeasy RNA purification kit (Qiagen); RLT lysis buffer was supplemented with β-mercaptoethanol (1%). Purified RNA was treated with DNase I (Invitrogen) before reverse transcription. cDNA was synthesized from RNA (1 μg) using iScriptTM cDNA Reverse Transcription kit (Bio-Rad) per manufacturer’s instructions. Real-time PCR was performed using the SYBR Green Master Mix kit and gene-specific primers. Quantitative PCR was performed on the ABI PRISM 7500 sequence detection system (Applied Biosystems, Foster City, CA). All reactions were performed in triplicates, and relative mRNA levels were calculated by the comparative threshold cycle method using GAPDH as an internal control. Oligonucleotide sequences are provided in Supplementary Table 1.

Rohatgi N., Zou W., Li Y., Cho K., Collins P.L., Tycksen E., Pandey G., DeSelm C.J., Patti G.J., Dey A, & Teitelbaum S.L. (2023). BAP1 promotes osteoclast function by metabolic reprogramming. Nature Communications, 14, 5923.

+ Open protocol

+ Expand

Quantitative mRNA Expression Analysis

Check if the same lab product or an alternative is used in the 5 most similar protocols

mRNA was purified from ~1.0 × 10⁶ cells using RNeasy RNA purification kit (Qiagen), and cDNA was produced using High Capacity cDNA Reverse Transcription kit (Applied Biosystems) according to the manufacturer protocol. qPCR reagent mixture was prepared by adding 10 ng of cDNA to TaqMan Fast Advanced Master mix (Thermo Fisher Scientific) and the corresponding TaqMan probes (FAM), which was then quantified by QuantStudio 7 Flex Real-Time PCR System (Thermo Fisher Scientific). GAPDH and 18S rRNA genes were used for data normalization.

An H., Ordureau A., Paulo J.A., Shoemaker C.J., Denic V, & Harper J.W. (2019). TEX264 is an ER-resident ATG8-interacting protein critical for endoplasmic reticulum remodeling during nutrient stress. Molecular cell, 74(5), 891-908.e10.

+ Open protocol

+ Expand

Transcriptome Analysis via Illumina Sequencing

Check if the same lab product or an alternative is used in the 5 most similar protocols

Each sample was ground in liquid nitrogen and total RNA was isolated using RNeasy RNA Purification Kit (Qiagen, Hilden) according to the manufacturer's protocol. To obtain complete gene expression information, a pooled RNA sample was prepared for transcriptome analysis. cDNA synthesis and libraries construction was performed following Illumina manufacturer's recommendations. In brief, poly (A)⁺ RNA was purified from 20µg total RNA using oligo (dT) magnetic beads and fragmented in presence of divalent cations under elevated temperature. Those fragmented poly (A)⁺ RNA were transcribed using random hexamer primers and this was followed by second strand cDNA synthesis using Rnase H and DNA polymerase I. The small fragments were purified using QiaQuick PCR Purification Kit. After that the purified fragments were washed with ethidium bromide buffer for the end-repair, poly (A)⁺ addition and ligation of sequencing adaptors. Suitable fragments (~200bp) were selected using agarose gel electrophoresis and amplified by PCR to construct a cDNA library.

Dong Y., Desneux N., Lei C, & Niu C. (2014). Transcriptome Characterization Analysis of Bactrocera minax and New Insights into Its Pupal Diapause Development with Gene Expression Analysis. International Journal of Biological Sciences, 10(9), 1051-1063.

+ Open protocol

+ Expand

Liver RNA Extraction and qRT-PCR Analysis

Check if the same lab product or an alternative is used in the 5 most similar protocols

Total RNA from fresh liver or cells was extracted using TRIzol (Invitrogen, Thermo Fisher Scientific) following RNA purification with RNeasy RNA purification kit and RNase free DNase digestion (QIAGEN, catalog 74104). Complementary DNA was synthesized from RNA (1 μg) using the iScript cDNA Reverse Transcription kit (Bio-Rad, catalog 1708890) according to the manufacturer’s instructions. Real-time PCR was performed using the SYBR Green Master Mix kit (Applied Biosystems, Thermo Fisher Scientific, catalog A25741) and the gene-specific primers listed in Table 1. GAPDH was used as a housekeeping gene. PCRs for each sample were performed with 7500 Fast Real-Time PCR System (Applied Biosystems, Thermo Fisher Scientific) using the comparative threshold cycle method for relative quantification.

Li Y., Su X., Rohatgi N., Zhang Y., Brestoff J.R., Shoghi K.I., Xu Y., Semenkovich C.F., Harris C.A., Peterson L.L., Weilbaecher K.N., Teitelbaum S.L, & Zou W. (2020). Hepatic lipids promote liver metastasis. JCI Insight, 5(17), e136215.

+ Open protocol

+ Expand

Quantification of CD22 Expression in CTCL

Cited 1 time

Check if the same lab product or an alternative is used in the 5 most similar protocols

Total cellular RNA was purified using the RNeasy RNA purification kit with the addition of on-column DNase digestion to avoid DNA contamination as described by the manufacturer (Qiagen; Hilden, Germany). Purified RNA was reverse transcribed using murine Moloney leukemia virus reverse transcriptase (M-MLV, Invitrogen) and oligo(dT) primer. The resulting cDNA was amplified by PCR using recombinant Taq DNA polymerase (no. M0267; New England Biolabs, Beverly, MA, USA) with the following primers: GAPDH forward: 5′-CCATGGAGAAGGCTGGGG, GAPDH reverse: 5′-CAAAGTTGTCATGGATGACC; CD22 preE5 forward: 5′-CTCCTAGAGGGGGTTCCAAT, CD22postE5 forward: 5′-GTCACCTCGACCTCCTTGAC, CD22E6 reverse: 5′-GGGAGTGACCTTGATCTCCA; CD22E4 forward: 5′-TTTTTGAGCACCCTGAAACC, CD22E5 reverse: 5′-CGGATACCCATAGCAGGAGA; CD22E11 forward: 5′-CATCCTCATCCTGGCAATCT, CD22E14 reverse: 5′-CTCTGCATCTCCAGTTCGTG. PCR program as follows: 94 ºC for 2 min, 32 cycles of 94 ºC for 15 s, at 56 ºC for 30 s, at 72 ºC for 30 s, and finally at 72 ºC for 10 min. The samples were analyzed in a 3 % agarose gel. The expression of CD22 in CTCL skin biopsy samples was analyzed using the above primers. The obtained expression results were standardized using genorm method utilizing ACTB, SDHA and B2M housekeeping genes [38 (link)].

Bagdonaite I., Wandall H.H., Litvinov I.V., Nastasi C., Becker J.C., Dabelsteen S., Geisler C., Bonefeld C.M., Zhang Q., Wasik M.A., Zhou Y., Sasseville D., Ødum N, & Woetmann A. (2015). Ectopic expression of a novel CD22 splice-variant regulates survival and proliferation in malignant T cells from cutaneous T cell lymphoma (CTCL) patients. Oncotarget, 6(16), 14374-14384.

+ Open protocol

+ Expand

Quantitative RT-PCR for Gene Expression

Check if the same lab product or an alternative is used in the 5 most similar protocols

Total RNA was extracted from cultured cells or zebrafish embryos with Trizol (Invitrogen) and purified with the RNeasy RNA purification kit (Qiagen). cDNA was synthesized with iScript cDNA synthesis kit (Bio-Rad), and quantitative RT-PCR was performed on StepOnePlus real time PCR system (Applied Biosystems) with SYBR Green probes (Applied Biosystems). Human or mouse hypoxanthine phosphoribosyltransferase 1 (HPRT1) gene or zebrafish beta-actin1 (actb1) gene was used as an internal reference respectively. Ratios of the expression level of each gene to the reference gene were then calculated with Microsoft Excel. Primers used in this study were listed in Supplementary Table S1.

Wang H., Lapek J., Fujimura K., Strnadel J., Liu B., Gonzalez D.J., Zhang W., Watson F., Yu V., Liu C., Melo C.M., Miller Y.I., Elliott K.C., Cheresh D.A, & Klemke R.L. (2018). Pseudopodium-enriched atypical kinase 1 mediates angiogenesis by modulating GATA2-dependent VEGFR2 transcription. Cell Discovery, 4, 26.

+ Open protocol

+ Expand

About PubCompare

Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.

We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.

However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.

Ready to get started?

Revolutionizing how scientists
search and build protocols!